Implantable biomaterials and medical devices are used in millions of procedures each year worldwide. However, in large number of patients, the implantation of these devices often leads to the development of a foreign body response (FBR), a chronic inflammatory condition that can ultimately lead to implant failure, which may cause harm to or death of the patient. The FBR consists of persistent inflammation coupled with fibrous encapsulation around the implant. There are no effective medical treatments. Hallmarks of the FBR include activation of macrophages at the tissue-implant interface, formation of destructive foreign body giant cells (FBGCs), and development of fibrous tissue that encapsulates the implant. The overall goal of our research is to understand the molecular mechanisms of the fibrotic response. Activated macrophages are thought to orchestrate the FBR by secreting inflammatory mediators. Emerging data support a critical role for a mechanical signal, e.g., substrate stiffness, in macrophage activation. However, a critical gap in this field is that the identity of the plasma membrane mechanosensor by which the mechanical signal is transduced/maintained is not known, nor are the downstream consequences of mechano-receptor signaling on the FBR. These gaps pose a significant barrier to progress in the field. In recent, exciting preliminary data, we obtained evidence that TRPV4, an ion channel in the transient receptor potential vanilloid family, and which is a known mechanosensor, may be the mediator of FBR. Specifically, we found that: 1) Trpv4 deletion in mice prevented macrophage accumulation, FBGC formation, and collagen accumulation in a subcutaneous implantation model; 2) the severity of the in vivo macrophage accumulation at the tissue-implant interface was dependent on the stiffness of the implant, and 3) genetic ablation or pharmacologic antagonism of TRPV4 blocked macrophage adhesion and spreading on stiff matrix, interleukin-4-induced FBGC formation, and inflammatory gene expression in both human and mouse bone marrow derived macrophages. Our preliminary data indicated that TRPV4 activity (Ca2+ influx) was augmented in response to increased matrix stiffness, and suggested that the molecular pathway linking TRPV4 activity to the FBR involved a specific phosphoinositide 3-kinase (PI3K) isoform, PI3K-alpha. The objective of this proposal is to determine the role of TRPV4 in the FBR. Based on our preliminary data, our central hypothesis is that TRPV4 mediates the FBR to biomaterials by increasing macrophage activation and fibrogenesis in a manner dependent on substrate stiffness and PI3K-alpha. We will utilize innovative technologies, in vivo and in vitro model systems, and a recently identified small molecule TRPV4 inhibitor to test the hypothesis with two Specific Aims. In Specific Aim 1 we will test the hypothesis that TRPV4 is a necessary component of the FBR in vivo; and in Specific Aim 2 we will test the hypothesis that mechanosensing by TRPV4 is a key component of the molecular mechanism of regulation of biomaterial- induced macrophage activation and fibrogenesis. When completed, we expect that the results of this study will generate novel information and insight regarding the mechanisms mediating the FBR to biomaterials, and will potentially identify a targetable receptor/pathway for the amelioration of FBR.